PROJECT SUMMARY The major goal of this proposal is to address the dynamic role of TREM2-mediated microglial function in brain aging and during different stages of the pathological development of Alzheimer?s disease (AD). TREM2 is a microglial specific gene with several of its rare variants associated with AD risk. Despite some progress, the molecular pathobiology of TREM2 in particular the TREM2-R47H risk variant is still not clear. Studies examining the effects of loss of TREM2 function in mouse models support inconsistent conclusions; with TREM2 deficiency either reduces or enhances amyloid or tau pathology and associated toxicity depending on the stage of the pathological development or the specific mouse models. As such, TREM2-mediated microglial function likely has dynamic effects on amyloid and tau pathologies depending on pathological stages throughout AD progression. Further complicating the challenge of studying the impact of TREM2-R47H variant, a recent study revealed that introducing the R47H mutation into the mouse Trem2 gene locus leads to aberrant splicing and instability of its mRNA. To fill these gaps in knowledge and the lack of appropriate model systems, we have generated novel cell type-specific and inducible mouse models expressing human TREM2 or TREM2- R47H in microglia. To address human relevance and molecular mechanisms, we have also generated human induced pluripotent stem cell (iPSC) lines carrying TREM2 or TREM2-R47H. Thus, the major goal of this proposal is to examine the dynamic effects of human TREM2 and TREM2-R47H on microglial and neuronal functions in aging and AD; while in the process defines the underlying molecular pathways by targeted and non-targeted approaches. We hypothesize that TREM2-mediated microglial function is protective against the development of AD pathologies but can be detrimental when such pathologies are associated with synaptic loss and neurodegeneration. We also hypothesize that TREM2-R47H represents a loss-of-function in particular in microglia-mediated protection against AD-related pathways. We will test our hypothesis through three aims. In Aim 1, we plan to analyze the effects of TREM2 or TREM2-R47H upon injury paradigms and during aging in the absence of AD pathology. In Aim 2, we will examine the effects of TREM2 or TREM2-R47H on the metabolism, deposition, and toxicity of A? and tau at different stages of pathological development. In Aim 3, we plan to identify and validate the molecular pathways associated with TREM2 and TREM2-R47H using iPSC- derived microglia-like cells with or without integration into cerebral organoids. This innovative proposal will take advantage of our existing conditional mouse models and iPSC-derived cellular models combined with state-of- the-art technologies including in vivo microdialysis, two-photon microscopy and molecular profiling by single cell RNA-Seq. These efforts should collectively help to understand how TREM2 modulates microglial dynamic roles in aging and AD pathogenesis and how we can target these pathways to treat AD.